Cardiovascular diseases (CVD) are a major cause of morbidity and mortality in postmenopausal women. In contrast this disease is rare before menopause. It is now believed that the majority of beneficial effects of estrogen (E2) on CVD are due to direct action of estrogen on the arterial wall. The process of aging has been shown to be associated with increased vascular smooth muscle (VSM) cell proliferation in response to growth factors and vascular injury. Since the severity and incidence of atherosclerosis is increased in postmenopausal women with advancing age, and since E2 has been shown to inhibit VSM cell proliferation, we postulate that E2 may mediate atheroprotective effects by inhibiting VSM cell proliferation. The long-term goal of this study is to understand the molecular mechanisms by which estrogen mediates its atheroprotective effects. We propose to develop gene therapy protocols based on the constitutively active human estrogen receptor in order to inhibit increased VSM cell proliferation associated with aging in females. We hypothesize that constitutively active human estrogen receptor-alpha (hER- alpha) gene transfer will decrease VSM cell proliferation in response to growth factors and vascular injury. We will determine the effect of hER-alpha gene expression on PDGF-BB- stimulated proliferation of VSM cells isolated from 24 month old female Fisher 344 rats. Further we will determine whether this inhibition is result of increased growth inhibitory effects of TGF-beta1 and/or inhibition of the MAP kinase pathway and protooncogene expression. We will use the balloon catheter- injured rat carotid artery model to test the effect of constitutively active hER-alpha gene transfer on neointima formation in 24 month old female rats. Gene transfer into VSM cells will be carried out in vitro and in vivo using replication deficient adenovirus-vector carrying constitutively active hER- alpha gene. Adenoviral vector carrying the beta-galactosidase gene (lacZ) will be used as controls in all studies. These pilot studies will help me to initiate a new area of research to investigate both age-related changes in VSM cells at cellular, biochemical, and molecular level and the mechanism of action of estrogen in VSM cells in female rats. Insights into the mechanisms by which the hER-alpha regulate VSM cell proliferation will offer new and exciting targets for treatment of menopause- related CVD in women.